CMV Induced Retinitis: Epidemiology, Clinical Features and Morbidity
Human CMV (HCMV) is a β herpesvirus which contains a dsDNA genome encoding >200 proteins (Chee et al., 1990, Curr Top Microbiol Immunol 154:125-169). HCMV infection usually develops asymptomatic lifelong infection in 50%-90% of healthy individuals, but can cause severe clinical complications when reactivated in immunocompromised patients (Ljungman, 2002, J Infect Dis: 186 Suppl 1:S99-S109; Fishman, 2007, N Engl J Med 357:2601-2614). One of the most serious HCMV-associated diseases in severely immunocompromised patients is the HCMV retinitis that leads to progressive loss of vision and blindness (Conboy et al., 1987, J Pediatr 111:343-348; Jabs et al., 1989, Ophthalmology 96:1092-1099; Jacobson and Mills, 1988, Ann Intern Med 108:585-594; Egbert et al., 1980, Ann Intern Med 93: 664-670; Pollard et al., 1980, Ann Intern Med 93: 655-664).
The incidence of HCMV retinitis in AIDS patients in the United States before the advent of highly active antiretroviral therapy (HAART), was estimated at 30% (Hoover et al., 1996, Arch Ophthalmol 114: 821-827), which has decreased to <10% in the post HAART era (Sugar et al., 2012. Am J Ophthalmol 153:1016-1024 e5; Palella et al., 1998, N Engl J Med 338:853-860; Jacobson et al., 2000, Clin Infect Dis 30:231-233). In children with symptomatic HCMV infection, an incidence of 5%-30% has been reported (Fowler et al., 1992, N Engl J Med 326:663-667). In severely iatrogenically immunosuppressed adult bone marrow transplant (BMT) recipients, one study has reported to have 10 HCMV retinitis cases of 5721 during a 14-year follow-up (Crippa et al., 2001, Clin Infect Dis 32:214-219). In solid organ transplant (SOT) recipients, a review of several studies has reported a total of 14 cases among 12,653 patients (Egli et al., 2008, Transpl Infect Dis 10:27-43). The clinical course for CMV retinitis can be protracted with prolonged periods of quiescence followed by progression. In HIV patients prior to HAART, CMV retinitis was associated with high rates of visual impairment (up to 98/100 eye-years (EYs]) and blindness (up to 49/100 EYs) (Holbrook et al., 2003, Arch Ophthalmol 121:99-107). In the modern era, treatment with antiretroviral agents can lead to suppression of the HIV RNA circulating in the blood (HIV load), thereby lending to immune recovery manifested as an increase in CD4+ T cells. Despite this dramatic decrease in the incidence of CMV retinitis and the improved outcomes due to modern antiretroviral therapy, CMV retinitis and vision loss from CMV retinitis continue to occur (Jabs et al., 2013, Ophthalmology 120:1262-1270; Jabs et al., 2010, Ophthalmology 117:2152-2161 e1-2), with the most recently reported rate of 0.9/100 person-year (Jabs et al., 2015, Ophthalmology pii:S0161-6420(15)00175-X, published online Apr. 16, 2015).
HCMV retinitis is diagnosed by ophthalmologic examination. Classic ophthalmologic findings of HCMV retinitis include white areas of retinal necrosis with associated hemorrhage and minimal vitreous inflammation (Lin et al., 2002, Retina 22:268-277). The current standard treatment of CMV retinitis consists of intravenous antiviral agents such as Ganciclovir and Foscarnet which are given at induction dosing for 2 weeks followed by maintenance dose of oral (Valganciclovir) or IV therapy for several weeks based on detection of CMV DNA in the blood or ophthalmologic evaluation. In patients failing to respond to these agents, cidofovir can be effective in clearing viremia or inducing regression of disease. Timely institution of treatment is critical, and in such cases approximately 50-60% of the patients will have either improvement or stabilization in visual acuity, while 40% of the patients will have progressive decline in vision (Eid et al., 2008, Transpl Infect Dis 10:13-18). Responses vary between the groups of patients depending on the underlying disease and level of immune suppression. The treatment is discontinued once there is evidence of reconstitution of T cell immunity. For patients who have ongoing immune suppression such as solid organ transplant recipients or AIDS patients with mid to higher level viral loads, current antiviral therapies carry significant toxicities when administered for prolonged periods. Therefore, additional therapies are needed for patients failing to respond, or for those who have continuous ongoing immune suppression.
Early studies in experimental mouse models provided the first evidence for the protective effect of adoptively transferred virus specific CD8 T cells against lethal, multiple-organ CMV infection. These studies used as immunocompromised host, BALB/c mice treated with hematoablative total-body irradiation, followed by intravenous adoptive transfer of CMV-primed CD8+ T cells and intra-plantar infection with murine CMV (MCMV) (Reddehase et al., 1988, J Virol 62:1061-1065; Reddehase et al., 1987, J Virol 61:3102-3108). Several subsequent studies defined the epitope specificities involved in protection against murine CMV and the additive effects of adoptively transferred CD4+ T-cells.
In humans, Riddell et al. first demonstrated the efficacy of adoptively transferred CMV-specific CD8 T-cell clones derived from the transplant donor prophylactically administered to recipients of BMT (bone marrow transplant) at risk for CMV infection (Riddell et al., 1992, Science 257:238-241; Walter et al., 1995, N Engl J Med 333:1038-1044). The efficacy of donor derived CMV-specific T-cells for the treatment of CMV viremia and disease was subsequently demonstrated (Einsele et al., 2002, Blood 99:3916-3922; Feuchtinger et al., 2010, Blood 116:4360-4367; Koehne et al., 2015, Biol Blood Marrow Transplant pii: S1083-8791(15)00372-9, published online May 29, 2015; Peggs et al., 2003, Lancet 362:1375-1377). Importantly, these initial trials demonstrated that CMV-specific T-cells can effectively treat CNS infections like encephalitis (Einsele et al., 2002, Blood 99:3916-3922; Feuchtinger et al., 2010, Blood 116:4360-4367), suggesting that these T-cells can penetrate the blood brain barrier. Similarly, in the treatment of Epstein-Barr virus related lymphoproliferative diseases (EBV-LPDs) developing in BMT recipients, it has been previously shown that adoptively transferred transplant donor derived EBV-specific T-cells can cause complete regressions of CNS lymphomas, providing evidence that adoptively transferred T-cells can home to the CNS (central nervous system) (Doubrovina et al., 2012, Blood 119:2644-2656).
Further advancements in this field evolved to address specific limitations of this therapy that would limit broad application of this treatment such as; the lack of timely availability of donor derived virus specific T-cells and the inability to generate cells from seronegative and cord blood donors. To overcome this limitation, pre-generated third party donor derived virus specific T-cells could be readily available for treatment of serious viral infections in such patients. Several groups have demonstrated the safety and potential efficacy of third party donor derived virus specific T cells for the treatment of EBV (Epstein-Barr virus), CMV and adenovirus infections in BMT and SOT (solid organ transplant) recipients (Hague et al., 2007, Blood 110:1123-1131; Leen et al., 2013, Blood 121:5113-5123).
Retina as an Immune Privilege Site: Pathogenesis and Implications for Treatment
The term ‘immune-privileged site’ was created in the 1940s by Sir Peter Medawar (Medawar, 1948, Br J Exp Pathol 29:58-69). In 1977, Barker and Billingham used this term to express the exemption of sites (such as the brain, ovary, testis, pregnant uterus, placenta, eye and the hamster cheek pouch) from immune responses (Barker and Billingham, 1977, Adv Immunol 25:1-54). Similarly, pathogen-mediated ocular inflammation can be harmful to the eye. Since minor inflammation can result in impaired vision or even blindness, the eye is naturally designed as an immune privileged site where infections usually do not lead to destructive immune reactions (Griffith et al., 1995, Science, 270:1189-1192). The underlying mechanism has been hypothesized to involve Fas ligand (FasL)-mediated programmed cell death (also called apoptosis) of Fas (CD95)-expressing T cells when attracted to the infection sites (Griffith et al., 1995, Science, 270:1189-1192). In this case, activated T cells are eliminated through ligation of Fas by FasL and no serious immune reactions are induced. TGF β is another cytokine present in the eye that inhibits Th1 cytokine mediated tissue destruction (Gabrielian et al., 1994, Invest Ophthalmol Vis Sci 35:4253-4259). Thus, the damage to the eye is minimized. However, CMV infection of human eyes is shown to cause large-scaled cell death and tremendous visual dysfunction (Jabs et al., 1989, Ophthalmology 96:1092-1099; Jacobson and Mills, 1988, Ann Intern Med 108:585-594).
The retina is anatomically protected from invading pathogens or inflammatory cells by the inner and outer blood-retina barrier. The inner blood-retina barrier consists of microvascular endothelial cells and the outer blood-retina barrier consists of RPE (retinal pigment epithelium) cells. Both cell types form functional tight junctions and are responsible for selective transport of essential molecules and for keeping out unwanted pathogens or activated leukocytes. It has been suggested that, in CMV infection, the internal blood-retinal barrier is disrupted after primary CMV replication in endothelial cells, allowing CMV particles to reach retinal glial cells. Subsequently, CMV might spread towards the RPE (Rao et al., 1998, Trans Am Ophthalmol Soc 96:111-126). Although glial cells, microvascular endothelial cells and RPE cells are major targets of CMV infection in the eye, all 10 layers of the retina are sites of necrotic lesions (Rao et al., 1998, Trans Am Ophthalmol Soc 96:111-126; Pecorella et al., 2000, Br J Ophthalmol 84:1275-1281; Palestine et al., 1984, Ophthalmology 91:1092-1099).
These anatomic and physiological features of the retina are thought to contribute to the characteristic features of CMV retinitis that are distinct from CMV infections in other organs; such as the occurrence of retinitis later in the course of infection or immunosuppression, in the absence of CMV viremia, and as a paradoxical infection occurring despite reconstitution of CD4+ T-cell count (Song et al., 2002, Retina 22:262-267). The same anatomical features can also potentially limit the efficacy of systemically administered anti-viral agents. For example, systemic CMV infections can be successfully treated with the anticytomegalovirus drugs ganciclovir, foscarnet or cidofovir, except in cases infected with ganciclovir resistant CMV strains. However, in some cases of retinitis, disease progression has been observed despite continuous antiviral therapy and proven drug sensitivity of isolated virus strains, suggesting that adequate drug concentrations of systemically administered drugs may not be achieved in the eye. Protocols for intravitreal administration of ganciclovir and foscarnet have therefore been implemented for treatment of CMV retinitis.
T-cell Immunity and CMV Infection
The adaptive immune system, particularly CD8+ T cells, plays a key role in the control of acute viral infections, including CMV infection (Crough and Khanna, 2009, Clin Microbiol Rev 22:76-98, Table of Contents). Viral-specific effector CD8+ T cells exert their antiviral activities through the production of type 1 cytokines such as interferon γ (IFN-γ) and tumor necrosis factor α (TNF-α), as well as through their antigen specific cytolytic activity. In both mouse models and in humans, reconstitution of CMV-specific CD8+ and CD4+ T-cells is critical for the control of CMV viremia and infections in recipients of both bone marrow (Quinnan et al., 1982, N Engl J Med 307:7-13; Hakki et al., 2003, Blood 102:3060-3067; Podlech et al., 1998, J Gen Virol 79:2099-2104) and solid organ transplants (Kumar et al., 2009, Am J Transplant 9:1214-1222; Pipeling et al., 2011, J Infect Dis 204:1663-1671).
CMV retinitis also occurs in patients with compromised cellular immunity, and in a proportion of patients, development of retinitis is associated with prior or concurrent reactivation of CMV in blood as well as other CMV end organ disease such as colitis or pneumonitis (Eid et al., 2008, Transpl Infect Dis 10:13-18). However, the median time to onset of retinitis is 6-12 months after initiation of immunosuppression or after transplant (Fishman, 2007, N Engl J Med 357:2601-2614; Crippa et al., 2001, Clin Infect Dis 32:214-219), which is a later time when the full effect of immunosuppressive medications in recipients of solid organ transplants are well established. For HIV patients, the primary clinical parameters determining risk for development of CMV retinitis are the post-HAART HIV viral load and CD4+ T helper cell counts ≤100/μl (Lin et al., 2002, Retina 22:268-277). CMV retinitis in HIV-positive patients also occurs later than CMV-associated conditions in other organs (colon, lungs, liver) affected by CMV infection. Furthermore, the T-cell response may also contribute to ocular pathology induced by CMV, as reflected by the development of uveitis in HIV patients at the time of CD4 T-cell recovery following institution of HAART therapy.
Taken together, these findings suggest that the rarity and late incidence of chorioretinitis reflect the profound degree of T-cell deficiency together with the resistance created by an intact blood-retinal barrier.
Earlier studies in murine CMV retinitis models have suggested that adoptive transfer of CMV-specific CD8+ T-cells could be protective against CMV retinitis developing in immunocompromised mice (Bigger et al., 1999, Invest Ophthalmol Vis Sci 40:2608-2613; Lu et al., 1997, Invest Ophthalmol Vis Sci 38:301-310). These studies used immunocompromised mice (thymectomized and T-cell depleted BALB/c mice) in whom retinitis was artificially induced by injecting infectious virus directly into the eye via the superciliary route, and groups of animals were infused with murine CMV-specific T-cells or control T-cells 2 hours prior to injection of virus. Although this model yielded the phenotypic and pathologic characteristics of CMV induced retinitis, this model is highly non-physiological, and the results cannot be extrapolated to the human retinitis treatment for several reasons. (1) In contrast to the mouse model in which CMV is injected directly into the eye, the intact retinal barrier is normally difficult to disrupt, and retinal human CMV infection develops through the retinal endothelium and occurs several months after immunosuppression, (2) The T-cells were transferred at the time of introduction of the infection in the murine model, while in human patients developing CMV retinitis, CMV-specific T-cells would be deficient or at least non-functional for some time (detailed above), and lastly (3) CMV is highly species specific. Early studies after isolation of murine and human cytomegaloviruses had observed that MCMV could not be propagated in human tissue and HCMV did not replicate in murine cells (Weller, 1970, J Infect Dis 122:532-539). After further exploration, it has been generally accepted that the cytomegaloviruses are highly species specific: Each virus replicates only in cells of its own or closely related host species. Therefore, murine CMV (MCMV) is not analogous to human CMV. There are also significant differences in the clinical spectrum of MCMV infection in that transplacental infection does not occur with MCMV, and even artificial introduction of MCMV at early stages of embryonic development does not result in CNS infection in mice, while human congenital CMV causes significant neurological sequelae. MCMV has been extensively studied and investigated for the potential use of this virus as a stand-in for human CMV (HCMV) to develop a mouse model, primarily for preclinical studies for vaccine strategies against human CMV, specifically to prevent congenital CMV infections. However, a chief limitation of the MCMV model for testing vaccines against congenital CMV infection has been the inability of the virus to infect the fetus by transplacental route, suggesting that MCMV causes a different spectrum of disease than HCMV. Although the precise molecular/cellular basis for the species-specificity of CMV remains unknown, studies by Maul et al. and others have shown that there are specific differences in the genes encoding the immediate early 1, 2 and 3 proteins of the MCMV and HCMV viruses, which have differential effects on the host cell transcriptional repressor Fax as well histone deacetylase, thereby affecting their ability for host infection (Maul and Negorev, 2008, Med Microbiol Immunol 197:241-249). These differences may also affect the host immune response to the virus, rendering this model less applicable to evaluation of immunotherapies for human CMV.
In order to overcome the species specificity of CMV infection, investigators have used human tissue explants; fetal thymus/liver (Mocarski et al., 1993, Proc Natl Acad Sci USA 90:104-108; Wang et al., 2005, J Virol 79:2115-2123; Brown et al., 1995, J Infect Dis 171:1599-1603) or fetal retinal tissue implants (Bidanset et al., 2001, J Infect Dis 184:192-195) maintained in SCID/hu mice that could allow the inoculation and propagation of HCMV in human cells which could then be used to test antiviral drugs or other therapeutic interventions specific for HCMV. Although these models are somewhat useful in evaluation of antiviral drugs (Kern, 2006, Antiviral Res 71:164-171), there is less information about exploiting the SCID/hu implant model for evaluation of HCMV vaccines and immunotherapies. Khanna and colleagues studied vaccine responses in a small animal model, in which HLA-2 transgenic mice immunized with replication-deficient adenovirus vectors expressing HCMV epitopes were used as a way to overcome species specificity of CMV viruses (Zhong et al., 2008, PLoS One 3:e3256). This chimeric vaccine demonstrated strong HCMV-specific CD8+ and CD4+ T-cell responses, as well as virus-neutralizing antibody. Although not a true “HCMV challenge” in a heterologous animal model, these experiments nevertheless represent an innovative approach to overcoming the problem of species specificity in CMV vaccine models.
Thus far it is clear that CMV retinitis exclusively occurs in patients with deficient T-cell immunity. In bone marrow transplant recipients, it has been reported that adoptive transfer of transplant donor derived or third party donor derived CMV specific T-cells restores T-cell immunity against CMV infection of the CNS such as encephalitis (Koehne et al., 2015, Biol Blood Marrow Transplant S1083-8791(15)00372-9, published online May 29, 2015; Feuchtinger et al., 2010, Blood 116:4360-4367), and in some cases of CMV retinitis (Gupta et al., 2015, Ophthalmic Surg Lasers Imaging Retina 46:80-82). Since third party donor derived T-cells have a limited survival after infusion before they undergo immune rejection within the allogeneic recipient, this approach may be used for bridging anti-CMV T-cell immunity in recipients of bone marrow transplants until reconstitution of T-cell immunity. However, there are specific differences in host physiology and CMV infection between recipients of solid organ transplant (SOT) and HIV-infected patients on one hand, and bone marrow transplant recipients on the other hand, because of which data from BMT recipients cannot be extrapolated to these two groups of patients. First of all, in recipients of SOT, CMV reactivation and original infection occurs within the donor cells (Hammond et al., 2013, Transpl Infect Dis 15:163-170; Harvala et al., 2013, J Med Virol 85:893-898), while BMT recipients experience host CMV reactivation. The infusion of third party CMV specific T-cells in SOT would thus carry the potential risk of precipitating immune rejection of the transplanted tissue carrying the CMV infection. Furthermore, independent of other factors, the inflammatory response resulting from CMV infection in SOT recipients could also trigger a rejection episode by enhancing antigen presentation, thus potentially placing these patients at high risk of organ allograft rejection by this treatment approach. Secondly, SOT recipients and HIV-infected patients have an indefinite period of immunodeficiency because of immunosuppressive therapy to prevent rejection, or variable CD4 count recovery with or without HAART therapy, respectively. In contrast, BMT recipients have a finite period of immunodeficiency in the absence of GvHD (graft-versus-host disease). This would necessitate multiple ongoing doses of CMV specific T-cells to treat CMV infection in SOT and HIV-infected patients, which could potentially compound the risk of allograft rejection in SOT recipients and immune recovery uveitis in HIV-infected patients. The efficacy of T cell therapy could also potentially be compromised by the lack of adequate number of T-cell doses in SOT recipients and HIV-infected patients.
Therefore, while there is a need for additional therapies for the treatment of CMV retinitis in human patients who are infected with HIV or who have been solid organ transplant recipients, these previous studies mentioned above have limited applicability.
Citation of a reference herein shall not be construed as an admission that such is prior art to the present disclosure.